This research proposal will be concerned with two problems of brain lipid metabolism which are individual both as to the methodology employed and the compounds under investigation. The enzyme protein(s) responsible for catalyzing the non-energy dependent, Ca ion-stimulatable "base exchange" reaction and phospholipase "D" activity will be purified. These two activities may reside in two distinct proteins or alternatively in a single protein. Phospholipase D cleaves the phosphodiester bond present in phospholipid, releasing a nonphosphorylated water-soluble product and phosphatidic acid. A reversal of this hydrolytic activity could be responsible for the "base exchange" reaction. It has been speculated that the low affinity choline uptake system of synaptosomes and brain slices may be a reflection of this activity. We have recently described the "solubilization" of these activities from rat brain particles. Classical methods of column chromatography and affinity chromatography will be exploited for further purification. Their catalytic, immunological and physiological roles will be investigated. The sphingoglycolipids are essential constituents of neuronal and glial cells as well as the myelin sheath. A variety of neurological diseases including infantile Gaucher's, Niemann-Picks, Krabbe's, metachromatic leukodystrophy, the gangliosidoses are characterized by altered metabolism of a specific sphingolipid. Pathways for the biosynthesis of these compounds have been proposed as a result of in vitro studies. However, there is a lack of substantiating in vivo data supporting these postulated pathways. L-Serine-C14 will be administered in vivo to label the sphingosine moiety of the sphingolipids; N- acetylamannosamine-(H3) to label the sialic acid residue of the gangliosides; N-acetylgalactosamine to label the amino sugar both of gangliosides and their asialo derivatives as well as the sialic acid of the gangliosides. Animals will be sacrificed at various times and the labeled sphingolipids isolated. These investigations should assist in elucidating the mechanism of synthesis of these compounds.